Área Biofísica, Facultad de Ciencias Bioquímicas y Farmacéuticas, Suipacha 531, S2002LRK Rosario, Argentina.
Photochem Photobiol Sci. 2012 Jun;11(6):962-6. doi: 10.1039/c2pp05341a. Epub 2012 Jan 25.
We have previously reported that bovine serum albumin (BSA) and other proteins that do not contain prosthetic groups exhibited a weak light absorption in the visible, only detectable by pulsed laser-induced optoacoustic spectroscopy (LIOAS). Human serum albumin (HSA) exhibited signals 25% higher than those observed with BSA. Signals comparable to those obtained with BSA were observed with poly(L-Trp, L-Lys), poly(L-Trp, L-Arg) or poly(L-Trp, L-Orn) at pH 7.0. No signals were obtained when tryptophan was replaced by other amino acids or when free tryptophan or the tripeptide Lys-Trp-Lys was assayed (pH 7.0). Tryptophan in HCl 5 N produced LIOAS signals similar to those produced by tryptophan-containing copolymers. Moreover, the absorption peak could be observed in a UV-VIS spectrophotometer. Therefore, the LIOAS signals obtained with BSA, HSA, and tryptophan-containing random copolymers may be attributed to a new transition of the indole moiety of their tryptophan residues when "protonated". Tryptophan residues of proteins are known to participate in π-cation interactions, which are important in protein stability and function. As a matter of fact, HSA and BSA contain an internal tryptophan in close proximity to lysine and arginine residues and therefore suitable for π-cation interactions. The strength of this type of interaction strongly depends on distances and relative orientations of both amino acid residues. Accordingly, these interactions should be highly sensitive to conformational changes. Based on preliminary results that have shown that LIOAS signal at 532 nm depended on the aggregation state of BSA and/or on the oxidation state of its Cys-34, we postulate that the LIOAS signal observed with proteins and tryptophan-containing polypeptides are related to Trp-Lys or Trp-Arg interactions and that the intensity of the signal depends on the strength of such interactions.
我们之前曾报道过，牛血清白蛋白（BSA）和其他不含辅基的蛋白质在可见光区有较弱的光吸收，只有通过脉冲激光诱导光声光谱（LIOAS）才能检测到。人血清白蛋白（HSA）的信号比 BSA 观察到的信号高 25%。在 pH 值为 7.0 时，聚（L-色氨酸，L-赖氨酸）、聚（L-色氨酸，L-精氨酸）或聚（L-色氨酸，L-鸟氨酸）的信号与 BSA 观察到的信号相当。当色氨酸被其他氨基酸取代或当游离色氨酸或三肽 Lys-Trp-Lys 被测定时（pH 值为 7.0），则没有信号。HCl 5N 中的色氨酸产生的 LIOAS 信号与含色氨酸的共聚物产生的信号相似。此外，在紫外-可见分光光度计中可以观察到吸收峰。因此，BSA、HSA 和含色氨酸的随机共聚物获得的 LIOAS 信号可能归因于其色氨酸残基“质子化”时吲哚部分的新跃迁。众所周知，蛋白质中的色氨酸残基参与π-阳离子相互作用，这对蛋白质的稳定性和功能很重要。事实上，HSA 和 BSA 都含有一个靠近赖氨酸和精氨酸残基的内部色氨酸，因此适合于π-阳离子相互作用。这种相互作用的强度强烈依赖于两个氨基酸残基的距离和相对取向。因此，这些相互作用应该对构象变化非常敏感。基于已经表明 532nm 处的 LIOAS 信号依赖于 BSA 的聚集状态和/或其 Cys-34 的氧化状态的初步结果，我们假设用蛋白质和含色氨酸的多肽观察到的 LIOAS 信号与 Trp-Lys 或 Trp-Arg 相互作用有关，并且信号的强度取决于这种相互作用的强度。
